The function(s) of a purified calcium-binding protein of brain will be evaluated in the proposed research. This protein has been identified as a Ca 2 ion-dependent regulator (CDR) of brain cyclic nucleotide phosphodiesterase and adenylate cyclase. Consequently, primary emphasis will be placed on exploring the role of CDR and Ca 2 ion in the regulation of brain cyclic nucleotide metabolism. Additional regulatory roles for CDR will also be sought in alternate enzyme systems. The approach to be pursued includes the following areas of investigation: (1) the purification to homogeneity, physical and kinetic characterization, and cellular localization of the Ca 2 ion-dependent phosphodiesterase by immunohistochemical procedures; (2) the development of agents which are specific inhibitors or alternate activators of the Ca 2 ion-dependent phosphodiesterase and their utilization in intact cells to define the role of this enzyme; (3) a detailed examination of the role of CDR in the modulation of various forms (particulate, hormone-sensitive, detergent-dispersed) of adenylate cyclase to define its involvement in cellular cAMP formation; (4) a comprehensive examination of the Ca 2 ion metabolism of a clonal line of glial tumor cells (C-6) containing CDR, CDR-dependent phosphodiesterase and CDR-dependent adenylate cyclase to define the relationship(s) among Ca 2 ion, CDR and cyclic nucleotide metabolism; (5) an exploration of the possible involvement of CDR in the regulation of other Ca 2 ion-dependent and lipid-modulated enzyme systems of brain; and (6)a more detailed physical characterization of CDR.